The proposed study intends to elucidate signal transduction mechanisms that establish molecular hierarchies in response to hypoxic challenges in salivary epithelial cells. Our current studies on the characterization of salivary signal transduction pathways have led to the identification of a novel non-receptor tyrosine kinase, Etk/BMX, expressed in rat parotid glands and rat salivary Pa-4 epithelial cells. We demonstrated that the paracellular barrier function and hypoxia-response element (HRE)-dependent gene expression could be enhanced upon the activation of Etk in salivary Pa-4 cells. In this application, we will test the hypothesis that Etk modulates the interplay between tight junctional component(s) and its interacting protein(s) for the maintenance of epithelial integrity under hypoxic condition. We will also test the hypothesis that Etk serves as a "molecular switch" to activate the HRE-dependent transcription, thereby providing cells the capacity to survive prolonged hypoxia. The long-term goal of this research is to use Etk-mediated cellular and biochemical modulations and gene regulation in salivary Pa-4 cells as a paradigm to elucidate mechanistic hierarchies in which the activation of a signaling pathway directs the adaptive responses to environmental stress. The proposed study is of great significance as the dysregulation of signaling pathway and/or hypoxia responses is associated with many human diseases, such as ischemic reperfusion and salivary tumors. There are four Specific Aims in this application: (i) To establish the role of Etk-dependent phosphorylation in salivary epithelial tight junction assembly/disassembly under hypoxia; (ii) To establish the mechanism by which Etk promotes tight junction assembly and enhances salivary epithelial barrier functions under normoxia and hypoxia; (iii) To determine whether Etk activation is essential for modulating salivary epithelial hypoxia-induced gene expression; and (iv) To verify and characterize selected Etk activation-induced target genes, which mediate adaptive responses to prolonged hypoxic exposure in salivary epithelial cells. These results will provide a new dimension to the Etk-mediated signaling pathways and mechanisms in salivary epithelial adaptive responses to hypoxia. A better knowledge of the molecular basis of Etk-activation and its effect on cellular, biochemical and genomic events under (patho) physiological conditions is key to successful future tissue engineering and drug discovery.